G418 (Geneticin)

G418 (Geneticin) : selection antibiotic, endotoxin tested, sterile reagent

G418 also known as G-418, G418 sulfate or Geneticin is an aminoglycoside antibiotic produced by Micromonospora rhodorangea.

G418 blocks polypeptide synthesis by inhibiting the elongation step in both prokaryotic and eukaryotic cells.

Resistance to G418 is conferred by the Neomycin resistance gene (neo) from Tn5 encoding an aminoglycoside 3'-phosphotransferase, APH 3' II.

Selection in mammalian cells is usually achieved in 3 to 7 days with concentrations ranging from 400 to 1000 µg/ml.

Cells that are dividing are affected more rapidly than non-dividing cells.

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Specifications

Product concentration: 100 mg/ml

CAS number: 108321-42-2

Quality Control: Each lot is thoroughly tested to ensure the absence of lot-to-lot variation.

Purity: ≥ 90% (HPLC)

Endotoxin level: < 0.5 EU/mg

Physicochemical characterization: pH, appearance

Cell-culture tested: potency validated in G418-sensitive and G418-resistant mammalian cell lines

Non-cytotoxicity of trace contaminants: absence of long-term effects confirmed in G418-resistant cells

Formula: C₂₀H₄₀N₄O₁₀. 2H₂SO₄

Molecular weight: 692.7

Structure:

Contents

G418 is supplied as a sterile filtered solution at 100 mg/ml in HEPES buffer.

This product is available in three pack sizes:

• ant-gn-1: 10 x 1 ml (1 g)
• ant-gn-2: 20 x 1 ml (2 g) 
• ant-gn-5: 1 x 50 ml (5 g)

G418 is shipped at room temperature.

Upon receipt it should be stored at 4°C.

Details

WORKING CONCENTRATIONS

G418 is normally used at a concentration of 400 μg/ml. However, the optimal concentration needs to be determined for your cells.
Suggested concentrations of G418 for selection in some examples of mammalian cells are listed below:

References:

1. Arikawa K. et al., 2003. Ligand-dependent inhibition of B16 melanoma cell migration and invasion via endogenous S1P2 G protein-coupled receptor. Requirement of inhibition of cellular RAC activity. J Biol Chem. 278:32841-51.
2. DiLillo DJ. et al., 2010. B cells are required for optimal CD4+ and CD8+ T cell tumor immunity: therapeutic B cell depletion enhances B16 melanoma growth in mice. J Immunol. 184:4006-16.
3. Figueroa K. et al., 2009. Selectivity of agonists for the active state of M1 to M4 muscarinic receptor subtypes. J Pharmacol Exp Ther. 28:331-42.
4. Kwon JA. et al., 2005. Biological function of the vaccinia virus Z-DNA-binding protein E3L: gene transactivation and anti-apoptotic activity in HeLa cells.
5. Hermoso M. et al., 2004. Cell volume regulation in response to hypotonicity is impaired in HeLa cells expressing a protein kinase Calpha mutant lacking kinase activity. J Biol Chem. 279:17681-9.
6. Hennen S. et al., 2013. Decoding signaling and function of the orphan G protein-coupled receptor GPR17 with a small-molecule agonist. Sci. Signal., 6:ra93.
7. Chen Q. et al., 2003. Expression of Drosophila trehalose-phosphate synthase in HEK-293 cells increases hypoxia tolerance. J Biol Chem. 278:49113-8.
8. Tukhvatulin A. et al., 2013. Combined stimulation of Toll-like receptor 5 and NOD1 strongly potentiates activity of NF-κB, resulting in enhanced innate immune reactions and resistance to Salmonella enterica serovar Typhimurium infection. Infect Immun. 81:3855-64.
9. Maue AC. et al., 2013. The polysaccharide capsule of Campylobacter jejuni modulates the host immune response. Infect Immun. 81:665-72.

FAQs

Why does one of my bottles of G418 solidify when stored at -20°C, while another remains completely liquid and translucent?
We observe the same phenomenon internally. Even below 0°C, the freezing reaction of the solution does not start spontaneously as it needs a disturbance to start. This disturbance can be a vibration, a shock, etc. According to our scientists, we often observe differences in solidification between several tubes of G418, which can be explained due to a small shock to one tube and not the others.
Please note that as soon as solidification has occurred in the liquid, it behaves like a chain reaction and spreads quickly throughout the available volume of solution.

What is the preferred storage method for G418?
G418 is shipped at room temperature. Upon receipt, it should be stored at 4°C for short term storage or -20°C for long term storage. Avoid repeated freeze-thaw cycles. The expiry date is specified on the product label. Once opened it remains stable for 1 year at -20°C providing you avoid repeated freeze-thaw cycles, and it can be kept at 4°C for 3 months provided it is kept in sterile conditions.

What is the selection concentration of G418 in mammalian cells?
The working concentration of G418 for selection and maintenance of mammalian cell lines transfected with the neo gene varies depending on a multitude of factors, including cell type. Therefore, we recommend to determine the optimal concentration of antibiotic required to kill your host cell line by initially treating your cells with several concentrations (e.g. 100 μg/ml – 1 mg/ml). Please note: G418 is regularly used at 400 μg/ml.

My plasmid has the Neo gene. Can I use G418 to select for it in E. coli?
No. G418 can only be used in mammalian cells expressing the Neo gene. The appropriate antibiotic for selection for E. coli transformants expressing the Neo gene is Kanamycin.

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